The evolution and spread of antibiotic resistance among bacteria is a major public health problem today, especially in the hospital setting with the emergence of multidrug resistant strains. Intensive research efforts have led to the development of new antibiotics effective against these resistant strains. Nevertheless, through use, mechanisms of resistance to these drugs emerge and limit their efficacy.
In view of this phenomenon, antimicrobial peptides (AMP) appear very promising for the design of new therapeutic agents. Cationic antimicrobial peptides are thought to be one of the key components of the innate immune system of multicellular organisms, which provides first-line defense against pathogens. The interest of these peptides lies on the one hand in their very broad spectrum of activity, enabling in particular their use in the treatment of infections caused by multidrug resistant strains. Secondly, their mode of action is based on permeabilization or rapid fragmentation of the microorganism membrane and is therefore unlikely to lead to the development of resistance mechanisms.
In particular, AMP have attracted considerable interest as potential agents against bacterial biofilms. Biofilms are bacteria that stick together, forming a community, which is embedded within a self-produced matrix. Biofilm bacteria show much greater resistance to antibiotics than their free-living counterparts and are responsible for various pathological conditions that are difficult to treat, such as chronic infection of patients affected with cystic fibrosis, endocarditis, and cystitis, infections caused by indwelling medical devices and dental plaque formation involved in caries and periodontitis. Since biofilm resistance to antibiotics is mainly due to the slow growth rate and low metabolic activity of bacteria in such communities, the use of AMP appears to be an attractive therapeutic approach because, due to their mode of action, they have a high potential to act also on slow growing or even non-growing bacteria. Antimicrobial peptides have been identified in plants, insects, amphibians and mammals. Amphibian skin represents a major source of antimicrobial peptides and every species of frog possesses its specific peptide repertoire generally composed of 10 to 15 AMP.
Frogs of the Ranidae family are very numerous and this family currently includes 16 genera and 338 species. These frogs synthesize and secrete a remarkable diversity of AMP, which have been classified into 13 families (Conlon et al., 2008 and 2009). One such family, the temporins, comprises AMP of small size (generally between 10 and 14 residues), the sequences of which vary widely according to species. More than 100 members of the temporin family have been identified. These temporins have been isolated from several Rana species such as Rana temporaria (Simmaco et al., 1996), Rana esculenta (Simmaco et al., 1990), Rana japonica (Isaacson et al., 2002), Rana ornativentris (Kim et al., 2001) and Pelophylax (Rana) saharica (Abbassi et al., 2008; Abbassi et al., 2010; Abbassi et al., 2013).
Unlike the other 12 families of Ranidae peptides, the temporins lack the “Rana box” motif, a C-terminal heptapeptide domain cyclized by a disulfide bridge (Mangoni, 2006). Furthermore, the majority of temporins contain a single basic residue, which confers a net charge of +2 at physiological pH. Generally, the temporins are particularly active against Gram-positive bacteria and yeasts but they also exhibit antifungal properties (Rollins-Smith et al., 2003) and, for some, antiviral properties (Chinchar et al., 2004).
It was found that temporin-SHa isolated from the skin of the North African frog Pelophylax saharica exhibits antiparasitic activity against protozoa belonging to the genus Leishmania, which are the causal agents of leishmaniosis (Abbassi et al., 2008). Based on this finding, analogues of said temporin exhibiting improved antimicrobial activity were obtained by substitution of one or more amino acids of the polar face of the α helix by a basic amino acid (WO 2010/106293). However, their toxicity, and in particular their hemolytic activity, constitutes an obstacle to their therapeutic uses, in particular if they are to be administered systematically.
Therefore, there is still a great need for improved antimicrobial peptides exhibiting strong antimicrobial activity and greatly reduced toxicity against mammalian cells.